This invention relates to antibiotic 5-substituted-3-(2-aminoethylthio)-6-(1-hydroxyethyl)-7-oxo-1-azabicyclo(3 .2.0)hept-2-ene-2-carboxylic acids (I): ##STR2## Wherein R is substituted or unsubstituted: alkyl having 1-6 carbon atoms, trifluoromethyl, phenyl, phenylalkyl having 7-12 carbon atoms, cycloalkyl having 3-6 carbon atoms, cycloalkylalkyl having 3-6 ring carbon atoms, and 1-6 carbon atoms in the alkyl moiety, and heteroaryl having 4-6 atoms in the ring, one or more of which is selected from O, N or S; and wherein the substituent or substituents on the foregoing radicals are selected from: OH, COOH, Cl, F, Br, NH.sub.2, alkyl and alkoxyl having 1-3 carbon atoms.
This invention also relates to the pharmaceutically acceptable salt and ester derivatives of I; pharmaceutical compositions comprising I and such derivatives; and to methods of treatment comprising administering such compounds and compositions when an antibiotic effect is indicated.
There is a continuing need for new antibiotics, for unfortunately there is no static effectiveness of a given antibiotic because continued wide scale usage of any such antibiotic selectively gives rise to resistant strains of pathogens. In addition, the known antibiotics suffer from the disadvantage of being effective only against certain types of microorganisms. Accordingly, the search for new antibiotics continues.
Unexpectedly, it has been found that the compounds of the present invention are broad spectrum antibiotics, which are useful in animal and human therapy and in inanimate systems.
Thus, it is an object of the present invention to provide a novel class of antibiotics which are 5-substituted analogues of the antibiotic thienamycin. These antibiotics are active against a broad range of pathogens which representatively include both gram positive bacteria such as S. aureus, Strep. pyogenes and B. subtilis and gram negative bacteria such as E. coli, Proteus morganii, Serratia, Pseudomonas and Klebsiella. Further objects of this invention are to provide chemical processes for the preparation of such antibiotics and their non-toxic pharmaceutically acceptable salts; pharmaceutical compositions comprising such antibiotics; and to provide methods of treatment comprising administering such antibiotics and compositions when an antibiotic effect is indicated.
The compounds of the present invention (I, above) are conveniently prepared by the following scheme: ##STR3##
In words relative to the above diagram, the 4-substituted-4-(2-substituted-vinyl)azetidine-2-one, 4, starting material is prepared by reacting an R.sup.1 -oxy-3-substituted-butadiene, 1, with chlorosulfonylisocyanate 2. The reaction is conducted without solvent or may be run in solvent such as diethyl ether, ethyl acetate, chloroform, methylene chloride, or the like, at a temperature of from -78.degree. C. to 25.degree. C. for from a few minutes to 1 hour to provide 3. The radical R.sup.1 is an easily removable acyl blocking group such as an alkanoyl or aralkanoyl which bears no functional group or groups which might interfere with the desired course of reaction (1+2.fwdarw.3.fwdarw.4). Intermediate species 3 is converted to the sulfinamide by reduction which is then hydrolyzed to 4 at pH 6-8. Typically the reaction solution comprising 3 is contacted (5-30 minutes) with an aqueous solution (at 0.degree.-25.degree. C.) of a reducing agent such as sodium sulfite, thiophenol, or the like, at pH 6-8 to provide 4.
The reaction 4.fwdarw.5 is a reduction, and is preferably achieved by hydrogenation in a solvent such as ethyl acetate ether, dioxane, tetrahydrofuran (THF), ethanol or the like at 0.degree. to 25.degree. C. for from 5 minutes to 2 hours under 1 to 10 atmospheres of hydrogen in the presence of a hydrogenation catalyst such as platinum metal or oxide thereof such as 10% Pd/C or the like.
The de-blocking reaction 5.fwdarw.6 is usually desirable when R.sup.1 is acyl to permit the later alkylation, 7.fwdarw.8. The preferred de-blocking procedure is by alcoholysis wherein the solvent is a lower alkanol such as methanol, ethanol or the like in the presence of the corresponding alkali metal alkoxide, such as sodium methoxide. Typically, the reaction is conducted for from 5 minutes to 1 hour at a temperature of from -10.degree. to 25.degree. C.
Blocking groups R.sup.3 and R.sup.2 are established (6.fwdarw.7) to provide a suitably protected species for alkylation (7.fwdarw.8). There is no criticality in the choice of blocking groups, provided only that they do not interfere with the intended alkylation. R.sup.3 may be hydrogen, a triorganosilyl group such as trimethylsilyl or the like, or a cyclic ether such as 2-tetrahydropyranyl; R.sup.2 may also be a cyclic ether such as 2-tetrahydropyranyl; alternatively R.sup.3 and R.sup.2 may be joined together to form protected species such as 7a: ##STR4## For example, species such as 7a are conveniently prepared by treating 6 with 2,2-dimethoxypropane in the presence of a catalyst such as boron trifluoride etherate, toluene sulphonic acid, or the like in a solvent such as methylene chloride, ether, chloroform, dioxane or the like at a temperature of from -10.degree. C. to 35.degree. C. for from a few minutes to 1 hour.
The alkylation (7.fwdarw.8) is preferably conducted by treating 7 with a strong base such as lithium diisopropylamide, sodium amide, potassium hydride or the like in a solvent such as THF, glyme, ether, dimethylformamide (DMF), dimethylsulfoxide (DMSO) or the like at a temperature of from -78.degree. C. to 0.degree. C. The resulting anion is then treated with excess acetaldehyde to provide 8.
The reaction 8.fwdarw.9 establishes the blocking group R.sup.4 and is typically accomplished by treating 8 with a base such as an alkali metal hydroxide, lithium diisopropylamide,4-dimethylaminopyridine, or n-butyllithium in a solvent such as ether, THF, dioxane, DMF, DMSO or the like, followed by treatment with an acyl halide of choice such as alkanoyl, aralkanoyl or nuclear substituted aralkanoyl, or alkyl, aryl or aralkyl, substituted aralkyl or substituted aryl haloformate such as p-nitrobenzylchloroformate, o-nitrobenzylchloroformate, or the like, at a temperature of from -78.degree. C. to 25.degree. C. for from 1-24 hours.
The de-blocking reaction 9.fwdarw.10 is typically conducted by acid hydrolysis such as aqueous acetic acid at a temperature of from 25.degree. C. to 75.degree. C. for from 5 minutes to 3 hours.
The aldehyde intermediate 11 is prepared by treating 10 with an oxidizing agent such as CrO.sub.3.2(pyridine) in CH.sub.3 CN, 1:1 mixture of dimethylsulfoxide and acetic anhydride, cyclohexylcarbodiimide in DMSO or the like at a temperature of from 0.degree.-25.degree. C. for from 5 minutes to 1 hour. The resulting species 11 in a solvent such as acetonitrile, methylene chloride, chloroform or the like at a temperature of from -10.degree. to 25.degree. C. is treated with an excess of N-blocked cysteamine, HSCH.sub.2 --CH.sub.2 NHR.sup.5, in the presence of an acid catalyst such as boron trifluoride etherate, toluene sulphonic acid or the like to provide 12. Typically, the reaction requires from 1 to 60 minutes.
There is no criticality as to the identity of the N-protecting group, R.sup.5, or the cysteamine reagent and suitable groups are p-nitrobenzyloxycarbonyl, o-nitrobenzyloxycarbonyl, t-butyloxycarbonyl, 2,2,2-trichloroethoxycarbonyl, phthaloyl, or the like.
The vinyl sulphide 14 is obtained via intermediate 13 by treating 12 with a halogen such as chlorine or bromine (X=Cl or Br) in a solvent such as ether, methylene chloride, tetrahydrofuran, glyme or the like at a temperature of from -78.degree. to 30.degree. C. for from 1 to 30 minutes, followed immediately by treating with an olefin such as cyclohexene, isobutylene, or the like in the presence of base such as triethylamine, DBU, sodium hydride, or the like in a solvent such as DMF, glyme, THF, HMPA. The solution is held at -20.degree. to 25.degree. C. for from 1 to 8 hours to yield 14.
The vinyl sulphide species 14 is reacted with a diester of oxomalonic acid (or its monohydrate) to provide 15. There is no criticality as to the identity of the ester moiety, R.sup.6, of the oxomalonic acid. R.sup.6 may be a conventional, easily removable blocking group or it may be a pharmaceutically acceptable ester moiety. Suitable ester radicals R.sup.6 are p-nitrobenzyl, benzyl, o-nitrobenzyl, t-butyl, 2,2,2-trichloroethyl. The reaction 14.fwdarw.15 is typically conducted in a high boiling organic solvent such as benzene, toluene, cyclohexane, halo aromatic or the like at a temperature of from about 50.degree. C. to reflux for from 0.5 to 6 hours.
The halogenation reaction 15.fwdarw.16 is typically conducted in a solvent such as THF, glyme, ether, methylene chloride, chloroform or the like in the presence of a halogenating agent such as thionyl chloride, phosphorous pentachloride or the like in the presence of base such as pyridine at a temperature of from -20.degree. to 25.degree. C. for from 5 minutes to 3 hours. The selective reduction of 15.fwdarw.17 via 16 is completed by treating 16 with tributylphosphine, triphenylphosphine or the like in aqueous DMF or similar aqueous systems involving dioxane, THF, glyme, DMSO, or acetone in the presence of K.sub.2 HPO.sub.4 at a temperature of from about 0.degree.-50.degree. C. for from 10 minutes to 5 hours.
Species 17 is halogenated by the previous procedure (12.fwdarw.13), but omitting the addition of the cyclohexene or other olefin, to provide the dihalo species 18. Species 18 is treated with a base such as triethylamine, sodium hydride or potassium hydride in a solvent such as DMF, acetonitrile, methylene chloride, chloroform, glyme or the like at a temperature of from about -78.degree. to 25.degree. C. for 1 to 5 hours to provide 19. Species 19 is converted to 20 on treatment with a strong base such as 1,5-diazabicyclo[5.4.0]undec-5-ene (DBU), 1,5-diazabicyclo[3.4.0]non-5-ene(DBN), or the like in a solvent such as DMSO, acetone, chloroform, DMF, THF, glyme or the like or on treatment with AgF in pyridine at a temperature of from 0.degree.-40.degree. C. for from 1/4 to 24 hours. The reaction 20.fwdarw.21 is conducted by treating 20 with an aromatic base such as pyridine, s-collidine or lutidine, in the presence of a displacing agent such as lithium iodide, lithium bromide, sodium bromide, or the like at a temperature of from about 80.degree.-150.degree. C. for from 15 minutes to 2 hours. An aqueous work up of the resulting reaction mixture provides 21. Isomerization of the double bond 21.fwdarw.22 is accomplished by treating 21 in a solvent such as DMF, DMSO, ethyl ether, THF, glyme, methylene chloride with a strong base such as diisopropylamine, DBU, DBN, or the like at a temperature of from 0.degree. to about 25.degree. C. for from a few minutes to 2 hours or until equilibrium has been established as determined by examination of sample aliquots by ultraviolet absorption or by thin layer chromatography. The final reaction 22.fwdarw.I (by hydrogenolysis of the blocking groups) is accomplished by treating 22 in a solvent such as dioxane, ethanol, THF or the like or an aqueous mixture thereof in the presence of a Platinum metal catalyst such as Pd/C under a hydrogen pressure of from 1-4 atmospheres for from 0.5 to 8 hours at a temperature of from about 0.degree.-25.degree. C.
The above-described total synthesis may also advantageously start with a 4-substituted-4-vinyl azetidinone (23), below rather than the enol acylate azetidinone (4, above). The following scheme illustrates this 4-substituted-4-vinyl-azetidinone embodiment of the present invention; notice that it ties into the above scheme at species 14. ##STR5##
In words relative to the above reaction diagram, the 4-substituted-4-vinyl azetidinone 23 is silylated to provide the N-silyl species 24. The groups R' on the silyl radical are loweralkyl having from 1-6 carbon atoms especially preferred triorganosilyl groups are trimethylsilyl and t-butyl-dimethylsilyl. Typically the silylation (23-24) is achieved by treating 23 in a solvent such as DMF, DMSO, HMPA or the like with the silylating agent of choice, dimethyl t-butylsilyl chloride, and a base such as Et.sub.3 N, pyridine, N,N-dimethylaniline and the like at a temperature of from -10.degree. to 30.degree. C. for from 1 to 8 hours. Species 24 is alkylated to form 25 by treatment with acetaldehyde in the presence of base. This reaction 24.fwdarw.25 is conducted exactly as described above for the alkylation 7.fwdarw.8. The O-protecting group is established in the reaction 25.fwdarw.26. The protecting group R.sup.4 is as previously defined and the reaction 25.fwdarw.26 is exactly analogous to the above described reaction 8.fwdarw.9. The removal of the N-triorganosilyl group is accomplished in reaction 26.fwdarw.27 by mild acid catalyzed solvolysis. The halo sulfide species 28 is obtained from 27 by treating 27 in a solvent such as methylene chloride, THF, glyme, or the like with the reagent XSCH.sub.2 CH.sub.2 NHR.sup.5 wherein R.sup.5 has previously been defined and X is halogen such as chloro or bromo at a temperature of from -50.degree. C. to 50.degree. C. for from 1 to 16 hours. The final sulfide intermediate 14, which is common to the above illustrated scheme of total synthesis is obtained from 28 by elimination of HX on treatment of 28 with a base such as 1,5-diazabicyclo (5.4.0)undec-5-ene (DBU), 1,5-diazabicyclo[4.3.0]non-5-ene, (DBN), 1,4-diazabicyclo[2.2.2]octane, (DABCO), or silver fluoride in a solvent such as DMSO, pyridine, DMF, HMPA or the like at a temperature of from -20.degree. to 50.degree. C. for from 1/4 to 16 hours.
The products of this invention (I) form a wide variety of pharmacologically acceptable salts with inorganic and organic bases; these include, for example, metal salts derived from alkali or alkaline earth metal hydroxides, carbonates or bicarbonates and salts derived from primary, secondary or tertiary amines such as monoalkylamines, dialkylamines, trialkylamines, loweralkanolamines, di-loweralkanolamines, lower alkylenediamines, N,N-diaralkyl lower alkylenediamines, aralkylamines, amino substituted lower alkanols, N,N-di-lower alkylamino substituted lower alkanols, amino-, polyamino- and guanidino-substituted lower alkanoic acids and nitrogen-containing heterocyclic amines. Representative examples include salts derived from sodium hydroxide, sodium carbonate, sodium bicarbonate, potassium carbonate, potassium hydroxide, calcium carbonate, trimethylamine, triethylamine, piperidine, morpholine, quinine, lysine, protamine, arginine, procaine, ethanolamine, morphine, benzylamine, ethylenediamine, N,N'-dibenzylethylenediamine, diethanolamine, piperazine, dimethylaminoethanol, 2-amino-2-methyl-1-propanol, theophylline, N-methylglucamine and the like. Acid addition salts, e.g., with hydrochloric, tartaric, hydrobromic, sulfuric nitric, toluene-p-sulphonic and methane sulphonic acids may also be employed.
The salts can be mono-salts such as the monosodium salt obtained by treating one equivalent of sodium hydroxide with one equivalent of the product (I), also mixed di-salts. Such salts may be obtained by treating one equivalent of a base having a divalent cation, such as calcium hydroxide, with one equivalent of the product (I). The salts of this invention are pharmacologically acceptable non-toxic derivatives which can be used as the active ingredient in suitable unit-dosage pharmaceutical forms. Also, they may be combined with other drugs to provide compositions having a broad spectrum of activity.
The compounds of the present invention are valuable antimicrobial substances which are active against various gram-positive and gram-negative pathogens. Thus the free acid, free base, and especially the salts thereof such as amine and metal salts, particularly the alkali metal and alkaline earth metal salts, are useful bactericides and can be used for removing susceptible pathogens from dental and medical equipment, for separating microorganisms, and for therapeutic use in humans and animals. For this latter purpose pharmacologically acceptable salts with inorganic and organic bases such as those known in the art and used for the administration of penicillins and cephalosporins can be utilized. For example, salts such as alkali metal and alkaline earth metal salts, and primary, secondary and tertiary amine salts can be used for this purpose. These salts can be combined with pharmaceutically acceptable liquid and solid vehicles to form suitable dosage unit forms such as pills, tablets, capsules suppositories, syrups, elixirs and the like which can be prepared in accordance with procedures well known in this art.
The novel compounds are valuable antibiotics active against various gram-positive and gram-negative bacteria, and accordingly, find utility in human and veterinary medicine. The compounds of this invention can therefore be used as antibacterial drugs for treating infections caused by gram-positive or gram-negative bacteria, for example against Staphylococcus aureus, Escherichia coli, Klebsiella pneumoniae, Bacillus subtilis, Salmonella typhosa, Pseudomonas and Bacterium proteus. The antibacterials of the invention may further be utilized as additives to animal feedingstuffs, for preserving foodstuffs and disinfectants. For example, they may be employed in aqueous compositions in concentrations ranging from 0.1 to 100 parts of antibiotic per million parts of solution in order to destroy and inhibit the growth of harmful bacteria on medical and dental equipment and as bactericides in industrial application, for example in waterbased paints and in the white water of paper mills to inhibit the growth of harmful bacteria.
The products of this invention may be used alone or in combination as an active ingredient in any one of a variety of pharmaceutical preparations. These antibiotics and their corresponding salts may be employed in capsule form or as tablets, powders or liquid solutions or as suspensions or elixirs. They may be administered orally, intravenously or intramuscularly.
The compositions are preparably presented in a form suitable for absorption by the gastro-intestinal tract. Tablets and capsules for oral administration may be in unit dose presentation form, and may contain conventional excipients such as binding agents, for example syrup, acacia, gelatin, sorbitol, tragacanth, or polyvinylpyrrolidone; fillers for example, lactose, sugar, maize-starch, calcium phosphate, sorbitol or glycine; lubricants, for example, magnesium stearate, talc, polyethylene glycol, silica; disintegrants, for example, potato starch or acceptable wetting agents such as sodium lauryl sulphate. The tablets may be coated according to methods well known in the art. Oral liquid preparations may be in the form of aqueous or oily suspension, solution, emulsions, syrups, elixirs, etc. or may be presented as a dry product, for reconstitution with water or other suitable vehicles before use. Such liquid preparations may contain conventional additives such as suspending agents, for example, sorbitol syrup, methyl cellulose, glucose/sugar syrup, gelatin, hydroxyethylcellulose, carboxymethyl cellulose, aluminum stearate gel or hydrogenated edible oils, for example almond oil, fractionated coconut oil, oily esters, propylene glycol, or ethyl alcohol; preservatives, for example methyl or propyl p-hydroxybenzoates or sorbic acid. Suppositories will contain conventional suppository bases, e.g., cocoa butter of other glyceride.
Compositions for injection may be presented in unit dose form in ampules, or in multidose containers with an added perservative. The compositions may take such forms as suspensions, solutions, or emulsions in oily or aqueous vehicles, and may contain formulatory agents such as suspending, stabilizing and/or dispersing agents. Alternatively, the active ingredient may be in powder form for reconstitution with a suitable vehicle, e.g., sterile, pyrogen-free water, before use.
The compositions may also be prepared in suitable forms for absorption through the mucous membranes of the nose and throat or bronchial tissues and may conveniently take the form of powder or liquids sprays or inhalants, lozenges, throat paints, etc. For medication of the eyes or ears, the preparations may be presented as individual capsules, in liquid or semi-solid form, or may be used as drops etc. Topical applications may be formulated in hydrophobic or hydrophilic bases as ointments, creams, lotions, paints, powders, etc.
Also, in addition to a carrier, the instant compositions may include other ingredients such as stabilizers, binders, antioxidants, preservatives, lubricators, suspending agents, viscosity agents or flavoring agents and the like. In addition, there may also be included in the composition other active ingredients to provide a broader spectrum of antibiotic activity.
For veterinary medicine the composition may, for example, be formulated as an intramammary preparation in either long acting or quick-release bases.
The dosage to be administered depends to a large extent upon the condition of the subject being treated and the weight of the host, the route and frequency of adminstration, the parenteral route being preferred for generalized infections and the oral route for intestinal infections. In general, a daily oral dosage consists of from about 5 to about 600 mg. of active ingredient per kg. of body weight of the subject in one or more applications per day. A preferred daily dosage for adult humans lies in the range of from about 15 to 240 mg. of active ingredient per kg. of body weight.
The instant compositions may be administered in several unit dosage forms as, for example, in solid or liquid orally ingestible dosage form. The compositions per unit dosage, whether liquid or solid may contain from 0.1% to 99% of active material, the preferred range being from about 10-60%. The compositions will generally contain from about 15 mg. to about 1500 mg. of the active ingredient; however, in general, it is preferable to employ a dosage amount in the range of from about 250 mg. of 1000 mg. In parenteral administration the unit dosage is usually the pure compound in a slightly acidified sterile water solution or in the form of a soluble powder intended for solution.